ABCMdb

PMID: 23702584

Zhou Y, Jiang Q, Takahagi S, Shao C, Uitto J
Premature termination codon read-through in the ABCC6 gene: potential treatment for pseudoxanthoma elasticum.
J Invest Dermatol. 2013 Dec;133(12):2672-7. doi: 10.1038/jid.2013.234. Epub 2013 May 23., [PubMed]

Sentences

No. Mutations Sentence Comment

4 ABCC6 p.Arg1141* Considering the redundancy of the genetic code, it was postulated that in case of the most common recurrent nonsense mutation, p.R1141X, the read-through may result in substitution of the arginine 1,141 by glycine, tryptophan, or cysteine. Login to comment 20 ABCC6 p.Arg1141* Received 1 March 2013; revised 17 April 2013; accepted 27 April 2013; accepted article preview online 23 May 2013; published online 27 June 2013 Abbreviations: mRNA, messenger RNA; PXE, pseudoxanthoma elasticum 2672 Journal of Investigative Dermatology (2013), Volume 133 & 2013 The Society for Investigative Dermatology recurrent nonsense mutation is p.R1141X, which accounts for B30% of all pathogenic PXE mutations in Caucasian patient populations. Login to comment 26 ABCC6 p.Arg1141* In this study, we focused on seven of them, including the most common stop codon mutation, p.R1141X, in which the arginine at position 1,141 (codon CGA) has been replaced by a stop codon (TGA) (Table 1). Login to comment 34 ABCC6 p.Arg1164* ABCC6 p.Arg1275* Testing of two mutant constructs, p.R1164X and p.R1275X, in the presence of varying concentrations of PTC124 indicated that the highest level of expression was noted with 5 mg ml 1 , as quantified by In-Cell ELISA (Figure 2). Login to comment 38 ABCC6 p.Arg1141* ABCC6 p.Arg1164* ABCC6 p.Arg518* ABCC6 p.Gln378* ABCC6 p.Arg1398* ABCC6 p.Arg1275* ABCC6 p.Gln1143* Different pseudoxanthoma elasticum (PXE)- associated nonsense mutations tested for PTC124 read-through Mutation Nucleotide sequence1 Location (exon) Mutation frequency (%)2 p.R1141X TGA-A 24 54 p.R1164X TGA-C 24 10 p.R518X TGA-G 12 1.2 p.R1398X TGA-G 29 1.2 p.Q378X TAG-A 9 o1 p.Q1143X TAG-G 24 o1 p.R1275X TGA-C 27 o1 1 The sequence depicts the stop codon (bold) followed by the nucleotide shown. Login to comment 42 ABCC6 p.Arg1141* ABCC6 p.Arg1275* An expression vector (p.CMV6-Entry) containing full-length wild-type human ABCC6 cDNA (a) or a corresponding mutant cDNA harboring p.R1141X premature termination codon containing a 30 -end sequence that encodes a DDK reporter peptide (b, c) or p.R1275X (d) was transfected into HEK293 cells in culture without (a, b) or with PTC124 (10mg ml 1 for 72hours) (c, d). Login to comment 50 ABCC6 p.Arg1141* In case of the p.R1141X mutation, considering the codon redundancy and possible combinations of the corrected mutations from the stop codon (TGA) to an amino acid, in addition to arginine (CGA), this position could be occupied either by cysteine (TGC) or tryptophan (TGG) or glycine (GGA). Login to comment 53 ABCC6 p.Arg1164* ABCC6 p.Arg1275* In this system, injection of 1to 4-cell-stage zebrafish embryos with a morpholino corresponding to the exon 7/intron 7 border of the 12 R1275X R1164X 10 8 6 4 2 0 0 1.2 2.5 5 10 PTC124 (&#b5;g ml-1) 20 40 Normalized OD at 460 nm Figure 2. Login to comment 54 ABCC6 p.Arg1164* ABCC6 p.Arg1275* Read-through efficiency of stop codon mutations p.R1275X and p.R1164X by PTC124 in different concentrations. Login to comment 71 ABCC6 p.Arg1141* In contrast, injection of human ABCC6 mRNA harboring the p.R1141X stop codon mutation failed to reverse the phenotype (Figure 5d and Table 2), suggesting that this mRNA rescue system can be used to check the pathogenicity of mutant human ABCC6 proteins, including those harboring the missense amino acids as a result of PTC124 read-through. Login to comment 87 ABCC6 p.Arg1141* However, injection of human ABCC6 mRNA, which harbors the p.R1141X mutation, does not result in phenotypic rescue (d). Login to comment 90 ABCC6 p.Arg1141* of embryos injected Lethality (%)2 Mo - 107 71.0 Scmo - 108 12.0 Mo&#fe; WT Arg 96 9.4 Mo&#fe; TGA STOP 102 85.3 Mo&#fe; TGG Trp 94 26.6 Mo&#fe; TGC Cys 134 7.5 Mo&#fe; GGA Gly 125 16.0 1 Zebrafish embryos were injected at day 0 with an abcc6a morpholino (Mo) alone or with human ABCC6 mRNA, either wild type (WT) harboring arginine at position 1,141, corresponding to the p.R1141X stop codon mutation (TGA), or containing a missense substitution of tryptophan (TGG), cysteine (TGC), or glycine (GGA) instead of arginine at position 1,141. Login to comment 93 ABCC6 p.Arg1141* ABCC6 with p.R1141X mutations in both alleles on the Abcc6 / background. Login to comment 95 ABCC6 p.Arg1141* It is conceivable that the presence of such an amino acid carried by a near-cognate transfer RNA (instead of the parent wild-type arginine in case of p.R1141X) could be pathogenic, essentially representing a missense mutation. Login to comment 132 ABCC6 p.Arg1141* ABCC6 p.Arg1141Cys ABCC6 p.Arg1141Trp Capped full-length human ABCC6 mRNA corresponding to wild-type, R1141X, or putative read-through substitutions of arginine1141 by cysteine, tryptophan, or glycine was transcribed from an expression vector pCMV-Tag4B using the T3 mMessage mMachine kit (Ambion, Austin, TX). Login to comment